Yak skin oligopeptide, preparation method and application thereof, and functional food or medicine

Abstract

The invention discloses a yak skin oligopeptide, a preparation method and application thereof, and a functional food or medicine, and belongs to the technical field of bioactive peptides, the yak skin oligopeptide comprises at least one of RDIFL, FRVP, RDILF and FDLRF oligopeptide, compared with other hypoglycemic peptides, the molecular weight is smaller, the yak skin oligopeptide can be directly absorbed by intestinal tracts, and the effect of delaying blood sugar rise is achieved. The oligopeptide RDIFL, the oligopeptide FRVP, the oligopeptide RDILF and the oligopeptide FDLRF have excellent inhibitory activity on alpha-glucosidase, the semi-inhibitory concentrations of the oligopeptide RDIFL, the oligopeptide FRVP, the oligopeptide RDILF and the oligopeptide FDLRF can reach 10.107 mmol / L, 13.063 mmol / L, 8.653 mmol / L and 12.369 mmol / L respectively, digestion and absorption of carbohydrates can be effectively delayed,

Description

Technical Field

[0001] The present invention relates to the technical field of bioactive peptides, in particular to a yak hide oligopeptide, a preparation method and application thereof, and a functional food or medicine. Background Art

[0002] Type 2 diabetes mellitus (T2DM), a disease characterized by persistent hyperglycemia and driven by a synergistic effect of genetic and environmental factors, has proven to be one of the most prevalent and fastest-growing diseases worldwide. It is characterized by insulin deficiency due to pancreatic β-cell dysfunction and insulin resistance. Key symptoms include elevated blood sugar and insulin resistance.

[0003] α-Glucosidase is an important enzyme in the body, primarily distributed within the brush border of intestinal cells. After ingestion, carbohydrates in food are first hydrolyzed into shorter polysaccharides by amylase in saliva. The partially hydrolyzed starch is then further converted by amylase secreted by the pancreas. When food reaches the small intestine, α-glucosidase hydrolyzes the α-1,4-glycosidic bonds of oligosaccharides and disaccharides, converting them into glucose for absorption. Therefore, limiting the catalytic activity of α-glucosidase using α-glucosidase inhibitors can effectively delay the digestion of dietary carbohydrates and thereby regulate postprandial hyperglycemia in patients with T2DM. However, clinically used α-glucosidase inhibitors, such as acarbose and voglibose, are expensive and have side effects such as abdominal pain and diarrhea. Therefore, the search for natural substances with α-glucosidase inhibitory activity has become a key area of ​​interest in diabetes research. Summary of the Invention

[0004] The purpose of the present invention is to provide a yak hide oligopeptide to address the technical defects in the prior art.

[0005] Another object of the present invention is to provide a method for preparing yak hide oligopeptides.

[0006] Another object of the present invention is to provide an application of yak hide oligopeptide.

[0007] Another object of the present invention is to provide a functional food.

[0008] Another object of the present invention is to provide a medicine.

[0009] The technical solution adopted to achieve the purpose of the present invention is: A yak hide oligopeptide, comprising at least one of RDIFL, FRVP, RDILF and FDLRF, wherein the amino acid sequence of RDIFL is arginine-aspartic acid-isoleucine-phenylalanine-leucine; the amino acid sequence of FRVP is phenylalanine-arginine-valine-proline; the amino acid sequence of RDILF is arginine-aspartic acid-isoleucine-leucine-phenylalanine; and the amino acid sequence of FDLRF is phenylalanine-aspartic acid-leucine-arginine-phenylalanine.

[0010] In the above technical solution, the yak skin oligopeptide is a bioactive peptide made from yak skin.

[0011] In the above technical solution, the RDIFL, FRVP, RDILF and FDLRF are all synthesized by solid phase synthesis.

[0012] Another aspect of the present invention also includes a method for preparing the yak hide oligopeptide, which is a solid phase synthesis method.

[0013] Another aspect of the present invention also includes a method for preparing the yak hide oligopeptide, comprising the following steps: Step 1, dissolving yak skin collagen peptide in water and then adding ethanol to obtain a mixture, allowing it to settle and centrifuging it to obtain the supernatant, concentrating it, and drying it to obtain crude yak skin peptide; Step 2: dissolving the crude yak skin peptide obtained in step 1 in ultrapure water, separating and purifying it by gel chromatography, using the aqueous solution as the eluent, collecting the effluent, concentrating it, and drying it to obtain the yak skin oligopeptide.

[0014] In the above technical solution, in step 1, the volume concentration of ethanol in the mixture is 75% to 85%.

[0015] In the above technical solution, in step 1, the static sedimentation time is 60 to 72 hours.

[0016] In the above technical solution, in step 1, the centrifugal speed is 6000-8000 r / min, and the centrifugal time is 15-20 min.

[0017] In the above technical solution, in step 1 and step 2, the drying is freeze-drying.

[0018] In the above technical solution, in step 2, the chromatographic column of the gel chromatography is TK-Col TA-GF30, and the detection wavelength of the ultraviolet detector is 220 nm.

[0019] In the above technical solution, in step 2, when gel chromatography is used for separation and purification, the effluent from 45 to 140 minutes is collected and concentrated.

[0020] Another aspect of the present invention also includes the use of the yak skin oligopeptide as an inhibitory peptide for α-glucosidase, and the molecular docking technology is used to verify the binding energy of the yak skin oligopeptide with α-glucosidase, and the binding energy is less than -8.9 kcal / mol.

[0021] Another aspect of the present invention also includes the use of the yak hide oligopeptide in the preparation of medicines for preventing and treating type 2 diabetes.

[0022] Another aspect of the present invention also includes a functional food comprising yak hide oligopeptide.

[0023] Another aspect of the present invention also includes a medicine comprising yak hide oligopeptide.

[0024] Compared with the prior art, the present invention has the following beneficial effects: 1. The four yak hide oligopeptides RDIFL, FRVP, RDILF and FDLRF of the present invention are all oligopeptides. Compared with other glucose-lowering peptides, they have a smaller molecular weight and can be directly absorbed by the intestines, playing a role in delaying the increase of blood sugar. 2. The oligopeptides RDIFL, FRVP, RDILF, and FDLRF of the present invention have excellent inhibitory activity against α-glucosidase, with half-inhibitory concentrations reaching 10.107 mmol / L, 13.063 mmol / L, 8.653 mmol / L, and 12.369 mmol / L, respectively. They can effectively delay the digestion and absorption of carbohydrates and reduce postprandial blood sugar. 3. This invention uses molecular docking technology to clarify the binding mechanism between yak skin oligopeptides and α-glucosidase, which is of great significance for the application of yak skin oligopeptides in food, medicine, and health products for improving type 2 diabetes; 4. The present invention broadens the diversified applications of yak hide, fully releases its economic and ecological value, and is of great significance for providing safer functional foods for the treatment or auxiliary treatment of type 2 diabetes. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 This is a gel chromatogram of the yak hide oligopeptides in Examples 2 to 4 of the present invention.

[0026] Figure 2 It is the inhibition rate of yak hide oligopeptides on α-glucosidase in Examples 2 to 4 of the present invention.

[0027] Figure 3 It is the high performance liquid chromatogram of the oligopeptide RDIFL of the present invention.

[0028] Figure 4 It is the mass spectrum of the oligopeptide RDIFL of the present invention.

[0029] Figure 5 It is the high performance liquid chromatogram of the oligopeptide FRVP of the present invention.

[0030] Figure 6 It is the mass spectrum of the oligopeptide FRVP of the present invention.

[0031] Figure 7 It is a high performance liquid chromatogram of the oligopeptide RDILF of the present invention.

[0032] Figure 8 It is the mass spectrum of the oligopeptide RDILF of the present invention.

[0033] Figure 9 It is a high performance liquid chromatogram of the oligopeptide FDLRF of the present invention.

[0034] Figure 10 It is the mass spectrum of the oligopeptide FDLRF of the present invention.

[0035] Figure 11 It is a global image of molecular docking and a molecular interaction diagram of the oligopeptide RDIFL of the present invention and α-glucosidase.

[0036] Figure 12 It is a global image of molecular docking and a molecular interaction diagram of the oligopeptide FRVP of the present invention and α-glucosidase.

[0037] Figure 13 It is a global image of molecular docking and a molecular interaction diagram of the oligopeptide RDILF of the present invention and α-glucosidase.

[0038] Figure 14 It is a global image of molecular docking and a molecular interaction diagram of the oligopeptide FDLRF of the present invention and α-glucosidase.

[0039] Figure 15 This is a graph showing changes in fasting blood glucose levels in each group of mice after continuous drug administration for 30 days in Application Example 4.

[0040] Figure 16 This is a graph showing changes in blood glucose levels of mice in each group at 0, 0.5, 1, 1.5, and 2 hours after oral administration of glucose in Application Example 4.

[0041] Figure 17 It is the area under the oral glucose tolerance curve of each group of mice in Application Example 4.

[0042] The α-glucosidase and p-nitrophenyl-α-D-pyranoglucopyranoside used in the following specific embodiments were purchased from Shanghai Yuanye Biotechnology Co., Ltd., DMSO (dimethyl sulfoxide) was purchased from Shanghai Haohong Biotechnology Co., Ltd., and the four oligopeptides RDIFL, FRVP, RDILF and FDLRF were synthesized by Hefei Sener Biotechnology Co., Ltd. through solid phase synthesis. DETAILED DESCRIPTION

[0043] The present invention will be further described in detail below with reference to specific embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the present invention.

[0044] Example 1 A yak hide oligopeptide comprises at least one of the four oligopeptides whose amino acid sequences are shown as SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3 and SEQ ID NO.4.

[0045] SEQ ID NO.1: RDIFL.

[0046] SEQ ID NO.2: FRVP.

[0047] SEQ ID NO.3: RDILF.

[0048] SEQ ID NO.4: FDLRF.

[0049] Among them, the molecular weight of RDIFL is 662.79 Da; the molecular weight of FRVP is 517.63 Da; the molecular weight of RDILF is 662.79 Da; and the molecular weight of FDLRF is 696.80 Da.

[0050] The four peptides, RDIFL, FRVP, RDILF, and FDLRF, are all oligopeptides with a small molecular weight and can be directly absorbed by the intestine. They specifically bind to α-glucosidase, exerting a role in delaying blood sugar elevation. Among them, the docking binding energy of RDIFL with α-glucosidase is -9.8kcal / mol; the docking binding energy of FRVP with α-glucosidase is -9.4kcal / mol; the docking binding energy of RDILF with α-glucosidase is -9.4kcal / mol; and the docking binding energy of FDLRF with α-glucosidase is -9.3kcal / mol. Therefore, all four oligopeptides have a good inhibitory effect on α-glucosidase and can prevent and treat type 2 diabetes.

[0051] Example 2 like Figure 1-Figure 2 As shown, this embodiment provides a method for preparing yak hide oligopeptide, comprising the following steps: Step 1, sugar removal: Dissolve yak skin collagen peptide in water and then add 4 times the volume of anhydrous ethanol to obtain a mixture, let it settle for 72 hours, centrifuge at 6000 rpm for 15 minutes, collect the supernatant, concentrate it by rotary evaporation at 55 rpm at 50°C, and freeze-dry to obtain yak skin crude peptide (powder); Step 2: Dissolve the crude yak skin peptide obtained in step 1 in ultrapure water and separate and purify it by TK-Col TA-GF30 gel chromatography. The loading concentration is 10 mg / ml, the loading volume is 5 ml, the detection wavelength of the UV detector is 220 nm, and the eluent is ultrapure water. Collect the effluent from tubes 1-2 (45-76 min), concentrate the effluent to an appropriate amount, and then freeze-dry it to obtain a yak skin oligopeptide with good solubility.

[0052] The yak skin oligopeptide obtained in this example was subjected to an α-glucosidase inhibitory activity assay, and the inhibition rate was found to be 22.13%.

[0053] Example 3 like Figure 1-Figure 2 As shown, a method for preparing yak skin oligopeptide is shown. The method of this embodiment is different from the method of Example 2 only in that the effluent from tubes 3-4 (77-110 minutes) is collected to prepare yak skin oligopeptide.

[0054] The yak skin oligopeptide obtained in this example was subjected to an α-glucosidase inhibitory activity assay, and the inhibition rate was determined to be 53.20%.

[0055] Example 4 like Figure 1-Figure 2 As shown, a method for preparing yak skin oligopeptide is shown. The method of this embodiment is different from the method of Example 2 only in that the effluent from the 5th tube (111-140 minutes) is collected to prepare the yak skin oligopeptide.

[0056] The yak skin oligopeptide obtained in this example was subjected to an α-glucosidase inhibitory activity assay, and the inhibition rate was found to be 33.86%.

[0057] Depend on Figure 1 As shown, F1, F2, and F3 are the yak skin oligopeptides obtained in Examples 2-4. It can be seen that the molecular weights of F1, F2, and F3 obtained by collecting the effluent at different times are different.

[0058] Depend on Figure 2It can be seen that after the yak skin oligopeptides (F1, F2 and F3) obtained in Examples 2-4 were subjected to α-glucosidase inhibitory activity assay, it was found that F2 had the strongest α-glucosidase activity, with an inhibition rate of 53.20%, and there was a significant difference between the inhibition rates of F1 and F3; followed by F3, with an inhibition rate of 33.86%, which was significantly different from the inhibition rate of F1.

[0059] Example 5 This embodiment provides an oligopeptide RDIFL having α-glucosidase inhibitory activity. The oligopeptide RDIFL was synthesized by conventional solid phase synthesis in the order of arginine-aspartic acid-isoleucine-phenylalanine-leucine. The oligopeptide RDIFL was detected by LC-MS / MS, and the obtained HPLC chromatogram was as shown in FIG. Figure 3 As shown in the mass spectrum Figure 4 The synthesized oligopeptide RDIFL has a molecular weight of 662.79 Da and a purity of >95%.

[0060] Example 6 This embodiment provides an oligopeptide FRVP having α-glucosidase inhibitory activity. The oligopeptide FRVP was synthesized in the order of phenylalanine-arginine-valine-proline using a conventional solid phase synthesis method. The oligopeptide FRVP was detected by LC-MS / MS, and the obtained HPLC chromatogram was as shown in FIG. Figure 5 As shown in the mass spectrum Figure 6 The synthesized oligopeptide FRVP has a molecular weight of 517.63 Da and a purity of >95%.

[0061] Example 7 This embodiment provides an oligopeptide RDILF having α-glucosidase inhibitory activity. The oligopeptide RDILF was synthesized by conventional solid phase synthesis in the order of arginine-aspartic acid-isoleucine-leucine-phenylalanine. It was detected by LC-MS / MS, and the obtained HPLC chromatogram was as shown in FIG. Figure 7 As shown in the mass spectrum Figure 8 The synthesized oligopeptide RDILF has a molecular weight of 662.79 Da and a purity of >95%.

[0062] Example 8 This embodiment provides an oligopeptide FDLRF having α-glucosidase inhibitory activity. The oligopeptide FDLRF was synthesized by conventional solid phase synthesis in the order of phenylalanine-aspartic acid-leucine-arginine-phenylalanine. It was detected by LC-MS / MS, and the obtained HPLC chromatogram was as shown in FIG. Figure 9 As shown in the mass spectrum Figure 10 The synthesized oligopeptide FDLRF has a molecular weight of 696.80 Da and a purity of >95%.

[0063] Application Example 1 The main components of the yak skin oligopeptides obtained in Examples 2 to 4 were determined by LC-MS / MS. The specific process of LC-MS / MS is as follows.

[0064] (1) After the yak skin oligopeptide was dissolved in the mobile phase as a sample, liquid phase separation was performed first, and then mass spectrometry detection was performed. The liquid phase used was a 0.1% v / v formic acid aqueous solution, and the liquid B was a 0.1% v / v formic acid acetonitrile aqueous solution (acetonitrile concentration was 84% ​​v / v). The liquid chromatography column was a PepMap RSLC C18 (75 um x 150 mm), equilibrated with 95% liquid A. The sample was loaded into Zorbax 300SB-C18 peptide traps (Agilent Technologies, Wilmington, DE) by an autosampler and then separated by a liquid chromatography column. The relevant liquid phase gradient was set as follows: from 0 min to 50 min, the linear gradient of liquid B was from 4% to 40%; from 50 min to 60 min, the linear gradient of liquid B was from 40% to 95%; from 60 min to 65 min, the liquid B was maintained at 95%.

[0065] (2) Yak hide oligopeptide samples were separated by capillary high performance liquid chromatography and then analyzed by mass spectrometry. The analysis time was 60 min. Detection method: positive ion. The mass-to-charge ratios of yak hide oligopeptide and its fragments were collected as follows: 10 fragment spectra (MS2 scan) were collected after each full scan.

[0066] (3) PEAKS software was used to search the database, and 5131 peptide sequences were obtained, including oligopeptides RDIFL, FRVP, RDILF, and FDLRF.

[0067] Application Example 2 The 5131 peptides obtained in Application Example 1 were screened for potential bioactive peptides, toxicity, water solubility, sensitization, overall average hydrophilicity and pharmacokinetics prediction, and finally the oligopeptides RDIFL, FRVP, RDILF and FDLRF were selected for verification based on the molecular docking binding energy.

[0068] like Figure 11-14 As shown, the safety and activity predictions were performed on the four oligopeptides RDIFL, FRVP, RDILF and FDLRF synthesized in Examples 5 to 8, respectively.

[0069] ToxinPred and AllerCatPro 2.0 predicted toxicity and allergenicity respectively. RDIFL, FRVP, RDILF and FDLRF were predicted to be non-toxic and non-allergenic.

[0070] The pharmacokinetic evaluation of the peptides showed that RDIFL, FRVP, RDILF and FDLRF had good absorption capacity in human intestine.

[0071] The 3D structure of α-glucosidase (PDB ID: 3A4A) was downloaded from the PDB database and used as the docking receptor. Pymol was used for structural preprocessing to remove water, solvent molecules, other ligands, and unnecessary metal ions, and hydrogen was added to the protein receptor. The 3D structure of the peptide was prepared using Novopro. Energy minimization was performed using the Calculations function and saved as an sdf file. This file was converted to a pbd file using Open Babel, and pdbqt files for the peptides were generated using Autodock Vina. Molecular docking was performed using AutoDock Vina 1.1.2.0 (the Scripps Research Institute, La Jolla, CA, USA), and visualization was performed using Pymol. The docking engine was AutoDock Vina, with the center coordinates (x, y, z) = (19.3, -3.3, 21.9) and the size set to (25.5 × 24.9 × 30.1).

[0072] The docking results are as follows Figures 11 to 14 As shown, arginine-aspartate-isoleucine-phenylalanine-leucine (RDIFL) forms 6 hydrogen bonds and 1 carbon-hydrogen bond with Ser240, Leu313, Arg315, Gln279, Glu277, and Asp215 residues of α-glucosidase, forming 1 pi-pi stacking, and there are 7 types of interactions; phenylalanine-arginine-valine-proline (FRVP) forms 8 hydrogen bonds with Asp352, Gln277, Gln279, Asp242, Lys156, Ser240, and Ser241 residues of α-glucosidase, forming 1 pi-pi stacking, and there are 3 types of interactions; Arginine-aspartate-isoleucine-leucine-phenylalanine (RDILF) forms four hydrogen bonds, three carbon-hydrogen bonds, and one Pi-Pi stacking with Arg315, Pro312, Tyr158, and Gln279 residues of α-glucosidase, and there are six types of interactions; phenylalanine-aspartate-leucine-arginine-phenylalanine (FDLRF) forms nine hydrogen bonds, one carbon-hydrogen bond, and one T-type Pi-Pi stacking with Asp242, Gln279, Asp307, Thr306, Gln353, Ser240, and Lys156 residues of α-glucosidase, and there are five types of interactions.

[0073] The binding energies of acarbose and four oligopeptides for α-glucosidase inhibition are shown in Table 1.

[0074] Table 1 Binding energy of oligopeptides for inhibition of α-glucosidase ; As can be seen from Table 1, compared with acarbose, the oligopeptide of the present invention has lower inhibitory binding energy with α-glucosidase and has good application prospects in preventing and treating type 2 diabetes.

[0075] Application Example 3 The half-inhibitory concentrations of the four oligopeptides RDIFL, FRVP, RDILF and FDLRF synthesized in Examples 5 to 8 were investigated as samples for α-glucosidase.

[0076] The in vitro inhibitory activity of α-glucosidase was determined using p-nitrophenyl-α-D-glucopyranoside (PNPG) as substrate.

[0077] Blank group: 120 uL of PBS and 50 uL (enzyme activity 5 U / ml) of α-glucosidase solution were mixed and incubated in a 37°C water bath for 20 min. Then, 25 uL of PNPG solution (8 mmol) was added and incubated for another 8 min before determination using a microplate reader.

[0078] Blank control group: add 120 uL of PBS and 25 uL of PNPG solution (8 mmol), incubate for 8 min, and then measure with a microplate reader.

[0079] Sample group: 120 μL of PBS was mixed with 75 μL of sample solution (oligopeptide concentration was 500 μM~20 mM, solvent was 0.1% DMSO) and 50 μL of α-glucosidase solution (enzyme activity was 5 U / ml) and incubated in a 37°C water bath for 20 min. Then, 25 μL of PNPG solution (8 mmol) was added and incubated for another 8 min before determination using a microplate reader.

[0080] Sample control group: 120 uL of PBS and 75 uL of sample solution (oligopeptide concentration is 500 μM~20 mM, solvent is 0.1% DMSO) were mixed and placed in a 37°C water bath for 20 min, then 25 uL of PNPG solution (8 mmol) was added and incubated for another 8 min before determination using a microplate reader.

[0081] The α-glucosidase inhibition rate was calculated according to the following formula.

[0082] α-glucosidase inhibition rate (%) = [1-(S-S0) / (C-C0)] × 100 Where: S is the absorbance of the sample group; S0 is the absorbance of the sample control group; C is the absorbance of the blank group; C0 is the absorbance of the blank control group.

[0083] Graphpad Prism 10 was used for fitting analysis and SPSS software was used for significance analysis. The analysis results are shown in Table 2. The IC values ​​of oligopeptides RDIFL, FRVP, RDILF, and FDLRF were 50 The values ​​were 10.107 mmol / L, 13.063 mmol / L, 8.653 mmol / L, and 12.369 mmol / L, respectively, indicating that the oligopeptides had a strong inhibitory ability against α-glucosidase.

[0084] Table 2 IC values ​​of active peptides for inhibition of α-glucosidase 50 value ; Note: a, b, c, d represent significant differences among different samples.

[0085] Application Example 4 like Figure 15-17 As shown, the blood sugar lowering effect of the yak hide crude peptide (powder) obtained in Example 2 was evaluated.

[0086] (1) Grouping and administration method: 40 SPF-grade 4-week-old male Kunming mice weighing 21-22 g were selected and adaptively fed for one week before the experiment. The 40 mice were randomly divided into 4 groups, namely normal control group (NC group), diabetic model group (DC group), positive control group (PC group), and Example 2 group (CP group). The mice in the normal control group (NC group) were fed with basic feed, and the other three groups were fed with high-sugar and high-fat feed for four consecutive weeks to induce diabetes. After four weeks, the three groups of mice were intraperitoneally injected with streptozotocin (STZ) dissolved in buffer for three consecutive days at a dose of 50 mg / kg; while the normal control group (NC group) was intraperitoneally injected with citrate buffer. One week later, the fasting blood glucose (FBG) levels of the mice in the diabetic model group (DC group), positive control group (PC group) and Example 2 group (CP group) exceeded 11.1 mmol / L -1 , the mice were identified as type 2 diabetic (T2DM) mice. The normal control group and the diabetic model group were then gavaged with 0.2 ml of sterile saline; the positive control group was gavaged with 200 mg / kg·BW of metformin. The Example 2 group was gavaged with the crude yak hide peptide powder from Example 2 dissolved in saline at a dose of 200 mg / kg·BW for 30 days.

[0087] (2) Determination of fasting blood glucose: Fasting blood glucose is a routine indicator for diagnosing type 2 diabetes. During the 30-day oral gavage period, the blood glucose levels of all mice were measured once a week at a fixed time using a handheld blood glucose meter. The mice were fasted but not watered for 8 hours before the measurement.

[0088] (3) Oral glucose tolerance test: Oral glucose tolerance test was performed three days before the end of the experiment. Mice were fasted for 8 h but not water, and then orally administered with glucose (2 g / kg body weight). Blood glucose levels were measured at 0, 0.5, 1, 1.5, and 2 h after the mice were induced using a handheld blood glucose meter.

[0089] (4) Data processing: SPSS 26 software was used for one-way analysis of variance. P < 0.05 was considered statistically significant.

[0090] Evaluation of hypoglycemic effect Figures 15-17 As shown. Figure 15 It can be seen that after oral administration of yak hide crude peptide, the fasting blood glucose level of the Example 2 group was reduced by 15.58% compared with the diabetic model group, indicating that yak hide crude peptide can inhibit the increase of blood glucose in diabetic mice.

[0091] Figure 16 The figure shows the changes in glucose tolerance of mice. It can be seen from the figure that compared with the diabetic model group, the blood glucose levels of the Example 2 group and the positive control group decreased faster. Figure 17 It can be seen that the Example 2 group significantly reduced the AUC value of T2DM mice within 120 minutes. The results show that yak skin crude peptide has a good regulatory effect on type 2 diabetes.

[0092] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A yak hide oligopeptide, characterized in that: The yak leather oligopeptide includes at least one of RDIFL, FRVP, RDILF and FDLRF, wherein the amino acid sequence of the RDIFL is arginine-aspartic acid-isoleucine-phenylalanine-leucine; the amino acid sequence of the FRVP is phenylalanine-arginine-valine-proline; the amino acid sequence of the RDILF is arginine-aspartic acid-isoleucine-leucine-phenylalanine; and the amino acid sequence of the FDLRF is phenylalanine-aspartic acid-leucine-arginine-phenylalanine.

2. The yak hide oligopeptide according to claim 1, characterized in that The yak hide oligopeptide is a bioactive peptide made from yak hide.

3. The method for preparing yak hide oligopeptide according to claim 1 or 2, wherein: The preparation method is a solid phase synthesis method.

4. The method for preparing yak hide oligopeptide according to claim 1 or 2, wherein: The following steps are involved: Step 1, dissolving yak skin collagen peptide in water and then adding ethanol to obtain a mixture, allowing it to settle and centrifuging it to obtain the supernatant, concentrating it, and drying it to obtain crude yak skin peptide; Step 2: dissolving the crude yak skin peptide obtained in step 1 in ultrapure water, separating and purifying it by gel chromatography, using the aqueous solution as the eluent, collecting the effluent, concentrating it, and drying it to obtain the yak skin oligopeptide.

5. The preparation method according to claim 4, characterized in that In step 1, the volume concentration of ethanol in the mixture is 75% to 85%; the static precipitation time is 60 to 72 hours; the centrifugal speed is 6000 to 8000 r / min, and the centrifugal time is 15 to 20 minutes; In step 1 and step 2, the drying is freeze-drying.

6. The preparation method according to claim 4, characterized in that In step 2, the gel chromatography column is TK-Col TA-GF30, and the detection wavelength of the ultraviolet detector is 220 nm; when gel chromatography is used for separation and purification, the effluent from 45 to 140 minutes is collected and concentrated.

7. Use of the yak hide oligopeptide according to claim 1 or 2 as an inhibitory peptide for α-glucosidase, characterized in that: The molecular docking technique was used to verify the binding energy between the yak skin oligopeptide and α-glucosidase, and the binding energy was less than -8.9 kcal / mol.

8. Use of the yak hide oligopeptide according to claim 1 or 2 in the preparation of medicines for preventing and treating type 2 diabetes.

9. A functional food, characterized in that The invention comprises the yak skin oligopeptide as claimed in claim 1 or 2.

10. A medicine, characterized in that The invention comprises the yak skin oligopeptide as claimed in claim 1 or 2.

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